Involvement of Mossy Cells in Sharp Wave-Ripple Activity In Vitro Aarti Swaminathan, Ines Wichert, Dietmar Schmitz, Nikolaus Maier  Cell Reports  Volume 23, Issue 9, Pages 2541-2549 (May 2018) DOI: 10.1016/j.celrep.2018.04.095 Copyright © 2018 The Author(s) Terms and Conditions

Cell Reports 2018 23, 2541-2549DOI: (10.1016/j.celrep.2018.04.095) Copyright © 2018 The Author(s) Terms and Conditions

Figure 1 Spontaneous MC Activity in Mouse Hippocampal Slices (A1) Reconstruction of a biocytin-filled MC. Dendrites in black, the axon in red. Below: the firing pattern during a 40 pA (1 s) current injection. (A2) Confocal image showing complex spines on the proximal dendrites. Arrowheads mark the axon. (A3) Magnification of the boxed areas in (A2) to visualize spines at higher resolution (arrows). (B) Example recordings (top) and raster plots of successive sweeps show varied discharge behaviors in MCs. (C) Histogram of overall spike rates. Inset: cumulative distribution of firing rates; red arrows indicate the cells shown in (B). (D1) Sketch to illustrate parallel LFP-MC recordings. (D2) Example showing the CA3 LFP and MC spiking. Asterisks indicate MC discharge concurrent with SWRs. Cell Reports 2018 23, 2541-2549DOI: (10.1016/j.celrep.2018.04.095) Copyright © 2018 The Author(s) Terms and Conditions

Figure 2 Analysis of MC Activity during CA3 SWRs (A1–A3) Discharge patterns of three MCs during CA3 SWRs. Successive sweeps (25 and 400 ms) centered to the SWR peak (average top). (B) Spiking of MCs within and outside SWR epochs was compared (Mann-Whitney U test). Left: display of p values (x axis, order of experiments). Red and orange dots indicate a significant increase (responding) or no significant increase (nonresponding) in spike rate during SWRs; dotted line, α = 0.001; arrows, cells shown in (A1–A3). Right: distribution of responding and nonresponding MCs. (C) Numbers of SWR epochs with MC spiking divided by total number of SWR epochs per experiment shown as percentages (x axis and color as in B). (D) Distribution of mean spike counts per SWR for responding cells (order as in B and C). (E) Left: correlation analysis of MC spiking and AP threshold (n = 21). Right: lower AP threshold in responding cells; error bars represent 10th and 90th percentiles. (F1) PETHs of responding MCs, 5 ms bin size. Top: grand average SWR. (F2) Average PETH after normalization (peak at 6.4 ms). Error bars represent SEM. Cell Reports 2018 23, 2541-2549DOI: (10.1016/j.celrep.2018.04.095) Copyright © 2018 The Author(s) Terms and Conditions

Figure 3 Timing of Ripple-Linked cPSCs in MCs (A1 and A2) SWRs in CA3c (top) linked with cEPSCs and cIPSCs in MCs (red and blue). (B1 and B2) Events marked in (A1) and (A2) at higher resolution; top to bottom: LFP SWR, filtered version, and cEPSC (red) and cIPSC (blue). Below: wavelet spectrograms of the cEPSC and cIPSC; warmer colors represent higher power. (C) Analysis of cPSC-to-ripple phases. Top: time points of steepest rising slopes in cPSC identified by their first derivatives (middle). The phases of these time points were determined with respect to the ripple (LFP) using its Hilbert phase (red dots, bottom). In total, 2,770 excitatory and 3,012 inhibitory slopes were analyzed. (D) Polar plots of average phases of excitatory (red) and inhibitory (blue) PSCs (slopes) of each cell with respect to CA3 ripples; black arrows: resultant phase vectors. (E1 and E2) Left, cross-correlation (CC) analysis of cEPSCs (E1) and cIPSCs (E2); single (gray) and averaged (red and blue) CC functions, aligned to peak of SPW envelope (top). Right: median SPW-cPSC time lags for cEPSCs (red) and cIPSCs (blue). Bottom: cIPSCs are delayed compared with cEPSCs. (F1–F3) Simultaneous PC-MC recordings during CA3c SWRs. (F1) Ripple peak-triggered averages of 100 SWRs (top) and their excitatory (left) and inhibitory (right) cPSCs (PC, green; MC, orange; reconstruction; F2). (F3) Median latencies for cEPSCs (red) and cIPSCs (blue). Inhibitory compared with excitatory latencies are consistently delayed in simultaneously recorded PCs and MCs (bottom). (E2) (bottom right) and (F3) (bottom): error bars represent 10th and 90th percentiles. Cell Reports 2018 23, 2541-2549DOI: (10.1016/j.celrep.2018.04.095) Copyright © 2018 The Author(s) Terms and Conditions

Figure 4 Ripple-Associated Functional Coupling of CA3 and GCs via MCs (A1) Reconstruction of simultaneously recorded MC and GC. (A2) Example sweeps, same experiment as (A1). Top: CA3 SWR and corresponding cEPSCs in MC (orange) and GC (blue). (A3) Ripple peak-triggered average of 100 SWRs (top) and corresponding cEPSCs (as A1 and A2). (B) Median latencies (16 MC-GC recordings; diamond, average). The error bar represents SEM. (C1–C3) Correlation of CA3c SWR and cEPSC amplitudes in PCs (C1), MCs (C2), and GCs (C3). Histograms of Pearson correlation coefficients for amplitudes of SWRs and cEPSCs in CA3 PCs (31 slices, 2,231 SWRs; C1), MCs (56 slices, 3,695 SWRs; C2), and GCs (38 slices, 2,326 SWRs; C3). Red lines, population medians. (D) Comparison of Fisher’s Z values. Dots, Z-transformed correlation coefficients from the histograms in (C1)–(C3). Error bars represent SEM. Cell Reports 2018 23, 2541-2549DOI: (10.1016/j.celrep.2018.04.095) Copyright © 2018 The Author(s) Terms and Conditions